S175: SINGLE-CELL ANALYSIS REVEALS SOMATIC TP53 MUTATIONS EMERGE IN EARLY PROGENITOR CELLS BUT BECOME ENRICHED IN THE ERYTHROID LINEAGE IN ERCC6L2 DISEASE
Authors: Laura Langohr, Ilse Kaaja, Suvi Douglas, Tuulia Räisänen, Sadiksha Adhikari, Markus Vähä-Koskela, Caroline Heckman, Jenni Lahtela, Ulla Wartiovaara-Kautto, Esa Pitkänen, Outi Kilpivaara
Journal: HemaSphere (2023)

Topic: 11. Bone marrow failure syndromes incl. PNH - Biology & Translational Research Background: ERCC6L2 disease is a recessively inherited bone marrow failure (BMF) characterized by high risk of progression to myelodysplastic syndrome (MDS) and further to acute myeloid leukemia (AML) with erythroid predominance (mostly AML M6) and poor prognosis (Douglas et al. 2019). ERCC6L2 patients acquire diverse somatic TP53 variants prior to the malignant transformation. First insights that a deficiency of ERCC6L2 in hematopoietic stem and progenitor cells leading to a delayed erythroid differentiation have been published recently (Armes et al. 2022). However, the effect of the somatic TP53 variants before and in the malignant transformation has not been addressed. Aims: The first aim of our study is to identify in which cell lineage and at which level of hematopoiesis TP53 variants occur. The second aim is to characterize the effect of the TP53 variants on the transcriptional landscape at single-cell level in ERCC6L2 disease, prior to and at progression to a hematological malignancy with erythroid characteristics. Methods: We integrated 3’ 10X single-cell transcriptome data from bone marrow samples from eight ERCC6L2 patients (seven BMFs and one MDS), 20 healthy donors (Oetjen et al.) and 21 AML patients using scVI (Lopez et al. 2018) and scANVI (Xu et al. 2021) with a hematological marker database (Triana et al. 2021). We identified TP53 variant status of single-cells from the same eight ERCC6L2 patient samples by targeted genotyping (Nam et al. 2019, Van Egeren et al. 2021) and a refined data analysis pipeline. Results: In total, we integrated data on 242 889 cells spanning 33 cell types. In line with clinical observations, the cell type proportions of ERCC6L2 patient samples resemble cell type proportions of healthy donor samples and not AML patient samples. We identified the TP53 variant status for 3 233 cells. First, we identified TP53 variants in myeloid, as well as in lymphoid cell lineages implying that the mutations have emerged in earlier common progenitor cells. Second, the TP53 mutation frequency was higher in erythroid cells than in other cell types (two-sided Fisher’s exact test, OR 1.50, 95% CI 1.27–1.79, adjusted p-value 1.60e-05). However, abundances of erythroid cells did not differ between ERCC6L2 patient cells compared to healthy donor cells (scCODA analysis, Büttner et al. 2021). Summary/Conclusion: Our single-cell transcriptome and TP53 targeted genotype data on ERCC6L2 patient samples reveal that somatic TP53 mutations emerge in early progenitor cells and become enriched in the erythroid lineage in ERCC6L2 disease already before malignant transformation. This is in line with the observations of prominent erythroid skewing in TP53-mutated (malignant) hematological diseases observed in ERCC6L2 patients. Our ongoing downstream data analyses focus on the characterisation of TP53 variants’ effects on the transcriptional landscape in ERCC6L2 disease. Overall, our study provides important insights of the disease dynamics leading to malignant transformation.Keywords: Bone marrow failure, TP53, Inherited disease, Big data